The conventional techniques of crop improvement in agriculture involve a search for strains of plants which exhibit new and useful characteristics, or to refine and improve on existing ones. The search has evolved from mere selection of a desirable parent plant to hybridization between parental strains which each exhibit desirable characteristics, finally, to crossbreeding between homozygous strains such that identical F.sub.1 progeny will be produced in each subsequent crossbreeding.
The conventional methods of maintaining genetic identity are well known and described in the literature. See, e.g., R. W. Allard "Principles of Plant Breeding," (John Wiley and Sons, Inc., 1960). The maintenance of purebred strains and the repeated crossbreeding to obtain F.sub.1 progeny are time consuming and labor intensive.
An additional limitation on the sexual reproduction of parental strains has been the low seed productivity per plant. This often results from low vigor which is manifested by heavily inbred strains. Finally, only a relatively limited number of purebred lines may be produced, and this results in a decreased pool of genetic characteristics available for selection.
It has been recognized that some of these difficulties may be overcome by vegetative propagation of the parental strain. See: W. C. Anderson and J. B. Carstens, "Tissue Culture Propagation of Broccoli, Brassica oleracea (Italica Group), for use in F.sub.1 Hybrid Seed Production," J. Amer. Soc. Hort. Sci., 102(1), pp. 69-73 (1977). This technique avoids the problem of the change in parental strain genetic characteristics through sexual reproduction. However, the sexual cross to produce F.sub.1 seed does not guarantee uniform progeny where there is chromosomal trait segregation in the parental strains.
It has been suggested that a desirable species may be propagated vegetatively with the somatic embryos or rooted plantlets being transferred to the field. However, this technique involves skilled labor in tissue culture laboratories, a transfer to a hothouse or nursery, and upon attaining sufficient acclimatization, a transplantation to the field. This procedure is costly and time consuming in comparison with the traditional methods of seeding.
To overcome some of these difficulties, the technique of fluid drilling has been developed. Fluid drilling methods have been used with pregerminated seed, e.g., D. Gray, "Comparison of Fluid Drilling and Conventional Establishment Techniques on Seedling Emergence and Crop Uniformity in Lettuce," J. Hort. Science, 52:23-30 (1978), and it has been suggested that fluid drilling may be adaptable to transfer somatic embryos directly to the field, e.g., D. A. Evans and W. R. Sharp, "Application of Tissue Culture Technology in the Agricultural Industry," in Application of Plant Cell and Tissue Culture to Agriculture and Industry, D. T. Tomes et al., eds. (University of Guelph Press, pp. 212-13, 1982). HoweVer, fluid drilling technology is capital intensive and requires the purchase of machinery and the development of new techniques in the agricultural community, which has been historically resistant to such change. Furthermore, fluid drilling does not allow for precision planting of seeds or somatic embryos.
To overcome the difficulties of fluid drilling, the creation of artificial seeds has been proposed in which somatic embryos are singly encapsulated in a hydrated gel consisting of 3.2% gel and 96.8% water (e.g., K. Redenbaugh, J. Nichol, M. E. Kossler, and B. Paasch, "Encapsulation of Somatic Embryos for Artificial Seed Production," In Vitro 20:256-257, 1984). The resultant hydrated capsule containing plant tissue can then be planted using traditional vacuum pick-up seed planters. However, the hydrated gel capsule contains encapsulated plant tissue that is also hydrated, consisting of a level of water equal to that of the gel capsule. When encapsulated, previously desiccated meristematic tissue, somatic embryos, or tissue-cultured plants would imbibe water to the level of the hydrated gel.
It has been suggested that somatic embryos be dried in a solution of polyethylene oxide. See: S. L. Kitto and Jules Janeck, "Production of Synthetic Seeds by Encapsulating Asexual Embryos of Carrot," J. Amer. Soc. Hort. Sci. 110(2):277-282 (1985). However, uncoated embryos did not survive desiccation. Furthermore, although coated clumps of embryos survived, no complete plants were produced.
This invention recognizes that the desiccation of plant tissue to a water content less than that of saturation of plant tissue may allow for more complete development and maturation, so that more vigorous, clonally uniform, and complete plants are formed. Furthermore, desiccated plant tissue may be more analogous to true seeds in terms of hydration level. Consequently, the desiccated plant tissue may be handled, stored, and treated in a manner closely analogous to true seeds.
Thus, an object of the invention is to provide a technique whereby cultured plant tissue may be insulated from harmful conditions.
Another object is to induce further maturation of the meristematic tissue, somatic embryos, or tissue-cultured plants, so that the plant tissue more readily, quickly, and uniformly forms a complete plant.
Still another object is to produce hardier and more developmentally complete meristematic tissue, somatic embryos, or tissue-cultured plants.
Another object is to control the developmental progression of plant tissue so that somaclonal and other variation are reduced or eliminated.
Yet another object is to reduce the water content of the plant tissue so that metabolism, DNA synthesis, and cell division are reduced and developmental arrest insues, allowing for an improved plant tissue with increased shelf life.
A further object is to desiccate the plant tissue so that DNA repair occurs.
Yet a further object is to desiccate the plant tissue to turn off development genes and turn on germination and growth genes.
Still another object is to desiccate the plant tissue so that hormone balance and plasma membrane integrity are achieved allowing for more complete maturation and whole plant recovery.
An additional object is to provide a clonal propagation system for genetically improved or transformed plants in which the modified genes or chromosomes are not stably integrated into the plant genome.
Another object of this invention is to decrease the time for raising a mature or vigorous seedling from meristematic tissues, somatic embryos or tissue-cultured plants.
Yet another object of the invention is to provide a medium to deliver the cultured plant tissue together with adjuvants facilitating seedling stand establishment.
A further object of the invention is to reduce the amount of handling between the development of the cultured plant tissue and its planting in the field.
A still further object of the invention is to reduce the need for special handling techniques and special technology during the development and growth of cultured plant tissue and, thus, overcome resistance to the introduction of new technology by adapting to existing methods of seed planting technology.
An additional object of the invention is to provide a large scale, economical method to clone superior plants or hybrid plants.